Clinical and immunological outcomes of HIV-exposed uninfected and HIV-unexposed uninfected children in the first 24 months of life in Western Kenya.

BACKGROUND: Previous studies show increased morbidity in children who are HIV-exposed but uninfected (HEU) compared to children who are HIV-unexposed uninfected (HUU). We sought to evaluate the effects of prenatal HIV exposure on clinical and immunological outcomes in the first 24 months of life. METHODS: Eighty-five HEU and 168 HUU children from Kenya were followed from birth to 24 months. All mothers living with HIV received combination antiretroviral therapy. Children who were HEU received standard-of-care cotrimoxazole prophylaxis through 18 months. Episodes of acute illness were identified through a combination of active and passive follow up. Trajectories of plasma cytokines, vaccine-specific antibodies, and antimalarial antibodies were examined. RESULTS: Children who were HEU and children who were HUU had similar growth curves. Children who were HEU had lower rates of malaria (rate ratio 0.54, 95% CI 0.38, 0.77) and respiratory illness (rate ratio 0.80, 95% CI 0.68, 0.93). Trajectories of plasma cytokines and vaccine-specific antibodies were similar in children who were HEU and HUU. There were subtle differences in antimalarial antibody dynamics, in which children who were HEU had overall lower antibody levels against five of the 14 malaria antigens tested. CONCLUSIONS: Children who were HEU and born to optimally treated mothers living with HIV had similar growth characteristics and immune profiles compared to children who were HUU. Children who were HEU had reduced risk for malaria and respiratory illness, which may be secondary to cotrimoxazole prophylaxis.

Hepatitis B virus infection status and associated factors among health care workers in selected hospitals in Kisumu County, Kenya: A cross-sectional study.

Poorly managed medical waste produced at the health facilities are potential source of infections including occupational exposure to Hepatitis B Virus (HBV). This study evaluated the prevalence of HBV infection among healthcare workers (HCWs) in Kisumu County. We determined prevalence of HBV infections among 192 HCWs from nine purposively selected high-patient volume public hospitals in Kisumu County. A structured questionnaire was administered, and 4.0 ml of venous blood sample collected for Hepatitis B surface antigen (HBsAg), hepatitis B surface antibody (anti-HBs) and total hepatitis B core antibody (anti-HBc) testing using enzyme immunoassay (EIA). Of 192 HCWs sampled, 52.1% were males and the median participants age was 34.4 years with interquartile range (IQR) of 11 (28-39) years. Most participants (44%) had worked for between 1-5 years. There was low HBV vaccine uptake with 35.9% completing the required 3 doses, while 40.6% had never been vaccinated. HBV prevalence was 18.8% (36/192), prevalence of past resolved infection was 25.5% (49/192), while 37.5% (72/192) of HCW had evidence of vaccine-derived immunity and 17.7% (34/192) were susceptible. HBV prevalence among HCW who had worked for less than one year and those who had never been vaccinated was 37.5% and 35.9% respectively. Significant risk of HBV lifetime exposure was noted among HCWs with one vaccine dose, those with no known exposure, while highest in those with knowledge on HBV transmission (aOR, 7.97; 95% CI, 2.10-153.3, p-value = 0.008). HCWs who had received ≥2 doses of HBV vaccine (aOR, 0.03; 95% CI, 0.01-0.10, p-value = <0.0001) had significant HBV protection. Duration of service was not associated with HBV among HCWs. HBV prevalence was high among HCWs from nine high patient volume public hospitals in Kisumu County. Efforts to strengthen HBV vaccination uptake and dose completion are needed to reduce HBV infections among HCWs.

Effects of Maternal HIV Infection on Early Kaposi Sarcoma-Associated Herpesvirus Seroconversion in a Kenyan Mother-Infant Cohort.

BACKGROUND: We identified whether maternal human immunodeficiency virus (HIV) infection during pregnancy affects transplacental transfer of Kaposi sarcoma-associated herpesvirus (KSHV)-specific antibodies and subsequent infant infection. METHODS: We followed pregnant Kenyan women through delivery and their infants until age 2 years. Children were classified as HIV-exposed uninfected (HEU) or HIV-unexposed uninfected (HUU) based on maternal HIV status. Maternal venous and cord blood at delivery and child venous blood every 6 months were tested for antibodies to 20 KSHV antigens by multiplex bead-based immunoassay. Multiple comparisons were adjusted using false discovery rate (FDR). RESULTS: Maternal HIV infection was significantly associated with decreased transplacental transfer of antibodies against all KSHV antigens and lower cord blood levels for 8 antigens at FDR P < .10. Neither birth to 6-month antibody level changes nor 6-month levels differed in HEU and HUU, except for ORF50. By age 24 months, 74% of children KSHV seroconverted but HEU and HUU did not differ in time to seroconversion nor 2-year seropositivity after adjustment for child malaria infection. CONCLUSIONS: Maternal HIV infection reduced a child's initial KSHV antibody levels but did not affect age of infection. Regardless of HIV exposure in utero, KSHV seroconversion in Kenyan children occurred early; associated factors must be identified.

Clinical and Immunological Outcomes of HIV-Exposed Uninfected and HIV-Unexposed Uninfected Children in the First 24 Months of Life in Western Kenya.

Background: Previous studies show increased morbidity in children who are HIV-exposed but uninfected (HEU) compared to children who are HIV-unexposed uninfected (HUU). We sought to evaluate the effects of prenatal HIV exposure on clinical and immunological outcomes in the first 24 months of life. Methods: Eighty-five HEU and 168 HUU children from Kenya were followed from birth to 24 months. All mothers with HIV received combination antiretroviral therapy. HEU children received standard-of-care cotrimoxazole prophylaxis through 18 months. Episodes of acute illness were identified through a combination of active and passive follow up. Trajectories of plasma cytokines, vaccine-specific antibodies, and antimalarial antibodies were examined. Results: HEU and HUU children had similar growth curves. HEU children had lower rates of malaria and respiratory illness. Trajectories of plasma cytokines and vaccine-specific antibodies were similar in HEU and HUU children. There were subtle differences in antimalarial antibody dynamics, in which HEU children had overall lower antibody levels against five of the 14 malaria antigens tested. Conclusions: HEU children born to optimally treated mothers living with HIV had similar growth characteristics and immune profiles compared to HUU children. HEU children had reduced risk for malaria and respiratory illness, which may be secondary to cotrimoxazole prophylaxis.

Cholera outbreak: antibiofilm activity, profiling of antibiotic-resistant genes and virulence factors of toxigenic Vibrio cholerae isolates reveals concerning traits.

Vibrio cholerae is a biofilm-forming pathogen with various virulence phenotypes and antimicrobial resistance traits. Phenotypic characteristics play a critical role in disease transmission and pathogenesis. The current study elucidated antibiofilm formation activity, profiled antibiotic-resistant genes and virulence factors of toxigenic Vibrio cholerae isolates from the cholera outbreak in Kisumu County, Kenya. Vibrio cholerae O1 isolates collected during the 2017 cholera outbreak in Kisumu County, Kenya, were utilized. Biofilm and virulence factors were profiled using standard procedures. The study confirmed 100 isolates as Vibrio cholerae , with 81 of them possessing cholera toxin gene (ctxA). Additionally, 99 of the isolates harboured the toxR gene. The study further revealed that 81 and 94 of the isolates harboured the class I integron (encoded by inDS gene) and integrating conjugative element (ICE), respectively. Antibiotic resistance assays confirmed tetracycline resistance genes as the most abundant (97 isolates). Among them were seven isolates resistant to commonly used antibiotics. The study further screened the isolates for antibiofilm formation using various antibiotics. Unlike the four strains (03/17-16, 02/17-09, 04/17-13), three of the strains (04/17-07, 06/17-14 and 05/17-03) did not form biofilms. Further, all the seven isolates that exhibited extensive antibiotic resistance produced haemolysin while 71.42%, 85.71 and 71.42 % of them produced protease, phospholipases and lipase, respectively. This study provides and in-depth understanding of essential features that were possibly responsible for V. cholerae outbreak. Understanding of these features is critical in the development of strategies to combat future outbreaks.

Molecular characterization of enterotoxigenic Escherichia coli toxins and colonization factors in children under five years with acute diarrhea attending Kisii Teaching and Referral Hospital, Kenya.

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of infectious diarrhea in children. There are no licensed vaccines against ETEC. This study aimed at characterizing Escherichia coli for ETEC enterotoxins and colonization factors from children < 5 years with acute diarrhea and had not taken antibiotics prior to seeking medical attention at the hospital. METHODS: A total of 225 randomly selected archived E. coli strains originally isolated from 225 children with acute diarrhea were cultured. DNA was extracted and screened by multiplex polymerase chain reaction (PCR) for three ETEC toxins. All positives were then screened for 11 colonization factors by PCR. RESULTS: Out of 225 E. coli strains tested, 23 (10.2%) were ETEC. Heat-stable toxin (ST) gene was detected in 16 (69.6%). ETEC isolates with heat-stable toxin of human origin (STh) and heat-stable toxin of porcine origin (STp) distributed as 11 (68.8%) and 5 (31.2%) respectively. Heat-labile toxin gene (LT) was detected in 5 (21.7%) of the ETEC isolates. Both ST and LT toxin genes were detected in 2 (8.7%) of the ETEC isolates. CF genes were detected in 14 (60.9%) ETEC strains with a majority having CS6 6 (42.9%) gene followed by a combination of CFA/I + CS21 gene detected in 3 (21.4%). CS14, CS3, CS7 and a combination of CS5 + CS6, CS2 + CS3 genes were detected equally in 1 (7.1%) ETEC isolate each. CFA/I, CS4, CS5, CS2, CS17/19, CS1/PCFO71 and CS21 genes tested were not detected. We did not detect CF genes in 9 (39.1%) ETEC isolates. More CFs were associated with ETEC strains with ST genes. CONCLUSION: ETEC strains with ST genes were the most common and had the most associated CFs. A majority of ETEC strains had CS6 gene. In 9 (39.1%) of the evaluated ETEC isolates, we did not detect an identifiable CF.

Severe acute respiratory syndrome coronavirus-2 antibody prevalence in people with and without HIV in rural Western Kenya, January to March 2020.

Among 582 participants in Western Kenya who were retrospectively tested from January through March 2020, 19 (3.3%) had detectable SARS-CoV-2 antibodies. The prevalence of detectable SARS-CoV-2 antibodies was similar between participants with and without HIV (3.1% vs. 4%, P = 0.68). One participant reported a cough in the preceding week but others denied symptoms. These may represent cross-reactivity or asymptomatic infections that predated the first reported COVID-19 cases in Kenya.

Malaria Is Associated With Kaposi Sarcoma-Associated Herpesvirus Seroconversion in a Cohort of Western Kenyan Children.

BACKGROUND: We aimed to determine whether Plasmodium falciparum infection affects age of Kaposi sarcoma-associated herpesvirus (KSHV) seroconversion in Kenyan children. METHODS: Kenyan children (n = 144) enrolled at age 1 month, from 2 sites with different levels of malaria transmission (stable/high vs unstable/low) were followed to age 24 months. Plasma was tested for KSHV antibodies using enzyme-linked immunosorbent assay (ELISA; K8.1 and LANA) and a multiplex bead-based assay (K8.1, K10.5, ORF38, ORF50, and LANA) and whole blood tested for P. falciparum DNA using quantitative PCR. Cox proportional hazards models were used to assess associations between P. falciparum DNA detection, malaria annualized rate (P. falciparum detections/person-years), and enrollment site (malaria-high vs malaria-low) with time to KSHV seroconversion. RESULTS: KSHV seroprevalence was 63% by age 2 years when assessed by multiplex assay. Children with P. falciparum were at increased hazards of earlier KSHV seroconversion and, among children with malaria, the hazard of becoming KSHV seropositive increased significantly with increasing malaria annualized rate. Children from the malaria-high transmission region had no significant difference in hazards of KSHV seroconversion at 12 months but were more likely to become KSHV seropositive by age 24 months. DISCUSSION: Malaria exposure increases the risk for KSHV seroconversion early in life.

Maternal HIV Infection as a Risk Factor for Primary Epstein-Barr Virus Infection in Kenyan Infants.

Human immunodeficiency virus (HIV) infection is known to be associated with EBV shedding in saliva suggesting an increased risk of EBV transmission to infants born to mothers with HIV at an earlier age. In this study we investigated (i) whether maternal HIV status was a risk factor for EBV in blood at delivery or for shedding in saliva and breast milk of 6- and 10-weeks post-partum mothers, (ii) if there was a difference in EBV strains shed between HIV+ and HIV- mothers, and (iii) if maternal HIV status was a determinant of EBV viral load in their infants. Samples were collected as part of a prospective cohort study that followed HIV-positive (HIV+) and HIV-negative (HIV-) pregnant women in Western Kenya through delivery and post-partum period. EBV viral load in blood was found to be significantly higher in mothers with HIV (p-value = 0.04). Additionally, a statistically significant difference was observed between EBV viral load in saliva samples and HIV status where HIV+ mothers had a higher EBV viral load in saliva at 6-weeks post-partum compared to HIV- mothers (p-value < 0.01). The difference in EBV shedding in breast milk was not found to be statistically significant. Furthermore, no difference in frequency of EBV strain was attributable to HIV- or HIV+ mothers. Interestingly, we found that infants born to HIV+ mothers had a higher EBV viral load at the time of their first EBV detection in blood than infants born to HIV- mothers and this was independent of age at detection. Overall, our study suggests that HIV infected mothers shed more virus in saliva than HIV-negative mothers and infants born to HIV+ mothers were at risk for loss of control of primary EBV infection as evidenced by higher EBV viral load following primary infection.

Benefits of enhanced infection prophylaxis at antiretroviral therapy initiation by cryptococcal antigen status.

OBJECTIVES: To assess baseline prevalence of cryptococcal antigen (CrAg) positivity; and its contribution to reductions in all-cause mortality, deaths from cryptococcus and unknown causes, and new cryptococcal disease in the REALITY trial. DESIGN: Retrospective CrAg testing of baseline and week-4 plasma samples in all 1805 African adults/children with CD4+ cell count less than 100 cells/µl starting antiretroviral therapy who were randomized to receive 12-week enhanced-prophylaxis (fluconazole 100 mg/day, azithromycin, isoniazid, cotrimoxazole) vs. standard-prophylaxis (cotrimoxazole). METHODS: Proportional hazards models were used to estimate the relative impact of enhanced-prophylaxis vs. standard-cotrimoxazole on all, cryptococcal and unknown deaths, and new cryptococcal disease, through 24 weeks, by baseline CrAg positivity. RESULTS: Excluding 24 (1.4%) participants with active/prior cryptococcal disease at enrolment (all treated for cryptococcal disease), 133/1781 (7.5%) participants were CrAg-positive. By 24 weeks, 105 standard-cotrimoxazole vs. 78 enhanced-prophylaxis participants died. Of nine standard-cotrimoxazole and three enhanced-prophylaxis cryptococcal deaths, seven and two, respectively, were CrAg-positive at baseline. Among deaths of unknown cause, only 1/46 standard-cotrimoxazole and 1/28 enhanced-prophylaxis were CrAg-positive at baseline. There was no evidence that relative reductions in new cryptococcal disease associated with enhanced-prophylaxis varied between baseline CrAg-positives [hazard-ratio = 0.36 (95% confidence interval 0.13-0.98), incidence 19.5 vs. 56.5/100 person-years] and CrAg-negatives [hazard-ratio = 0.33 (0.03-3.14), incidence 0.3 vs. 0.9/100 person-years; Pheterogeneity = 0.95]; nor for all deaths, cryptococcal deaths or unknown deaths (Pheterogeneity > 0.3). CONCLUSION: Relative reductions in cryptococcal disease/death did not depend on CrAg status. Deaths of unknown cause were unlikely to be cryptococcus-related; plausibly azithromycin contributed to their reduction. Findings support including 100 mg fluconazole in an enhanced-prophylaxis package at antiretroviral therapy initiation where CrAg screening is unavailable/impractical.

Malaria during pregnancy and transplacental transfer of Kaposi sarcoma-associated herpesvirus (KSHV) antibodies: a cohort study of Kenyan mother and child pairs.

BACKGROUND: Kaposi sarcoma-associated herpesvirus (KSHV) seroprevalence in sub-Saharan African children can range up to 50% by age 2 years but factors affecting early age of KSHV infection are not well understood. Malaria during pregnancy has been associated with hindered transplacental transfer of antibodies to several pathogens but whether it affects transplacental transfer of KSHV antibodies is unknown. We aimed to determine if in utero malaria exposure reduced the transfer of KSHV antibodies across the placenta. METHODS: A cohort study in Kisumu, Kenya enrolled pregnant women at their first antenatal clinic (ANC) visit and followed them through delivery. We included 70 KSHV-positive, HIV-negative mothers and their children. KSHV antibody levels were measured by ELISA (K8.1, ORF73) and multiplex assay (K8.1, ORF73, K10.5, ORF38, ORF50). Transplacental transfer of antibodies was measured by the cord to maternal blood ratio (CMR) of KSHV antibodies. Malaria during pregnancy was defined as detection of Plasmodium falciparum (Pf) DNA at any ANC visit or delivery. Among women with malaria during pregnancy, we examined time of last malaria infection prior to delivery (< 27 vs. 27+ weeks gestation) and malaria incidence rate (MIR) (episodes/100 person-weeks). RESULTS: KSHV seroprevalence (positive for K8.1 or ORF73 by ELISA) among pregnant women was 88%. Neither malaria during pregnancy, malaria infection timing, nor MIR were associated with maternal delivery KSHV antibody blood levels. Maternal delivery and cord blood KSHV antibody levels were highly correlated but these correlations did not differ by malaria during pregnancy. KSHV transplacental antibody transfer was not associated with malaria during pregnancy, malaria infection timing, nor MIR. CONCLUSIONS: Malaria during pregnancy does not appear to affect transfer of KSHV antibodies across the placenta.

Geographical distribution and antibiotics susceptibility patterns of toxigenic Vibrio cholerae isolates from Kisumu County, Kenya.

BACKGROUND: Multiple drug resistance has become a major threat to the treatment of cholera. Recent studies in Kenya have described the epidemiology, especially the risk factors, of cholera; however, there is little information on the phenotypic and drug susceptibility patterns of Vibrio cholerae (V. cholerae) in outbreaks that in the recent past have occurred in western Kenya. AIM: To characterise and determine the antibiotics' susceptibility profiling of toxigenic V. cholerae isolates from Kisumu County. SETTING: The project was conducted in Kisumu County, Kenya. METHODS: A total of 119 V. cholerae O1, biotype El Tor, isolates collected during 2017 cholera outbreak in Kisumu County were used for this study. The samples were cultured on thiosulphate-citrate-bile salts sucrose (TCBS) agar and biochemical tests were carried out using standard procedures. Susceptibility tests were conducted by using various conventional antibiotics against standard procedures. RESULTS: Of the 119 isolates, 101 were confirmed to be V. cholerae belonging to serotypes Inaba and Ogawa, with Inaba being the predominant serotype (73.95%). The isolates were susceptible to ciprofloxacin (100%), ofloxacin (100%), gentamycin (100%), doxycycline (99%), ceftriaxone (99%) and streptomycin (96.04%) antimicrobials, and resistant to erythromycin (53.47%), amoxicillin (64.4%), nalidixic acid (83.2%) and ampicillin (89.11%), with high resistance to cotrimoxazole (99%) and tetracycline (97%). CONCLUSION: Vibrio cholerae was resistant to multiple antibiotics, including those commonly used in the management of cholera. Taken together, there is a need to carry out regular surveillance on antimicrobial drug resistance during outbreaks.

Early cardiac dysfunction in children and young adults with perinatally acquired HIV.

OBJECTIVE: To define the prevalence of early cardiac dysfunction in children and young adults with perinatally acquired HIV and predictors of cardiac function. DESIGN: Cross-sectional design. METHODS: Early cardiac dysfunction was defined as left ventricular (LV) global longitudinal strain z-score less than -2 or myocardial performance index at least 0.5 with normal LV ejection fraction. Regression models were fitted to assess the relationship between measures of cardiac function and HIV RNA levels, clinical variables, and markers of inflammation. RESULTS: Six hundred and forty-three individuals (mean age 14.1 ±â€Š5.2 years) were enrolled. The average time on combination antiretroviral treatment was 6.8 ±â€Š3.6 years. Nearly 28% of individuals met criteria for early cardiac dysfunction. Individuals with early cardiac dysfunction were older (15.3 vs. 13.5 years, P < 0.001), had more frequently detectable HIV RNA (52.5 vs. 41.7%, P = 0.018), were more likely exposed to azidothymidine or zidovudine (ZDV) (55.6 vs. 41.2%, P = 0.002), and had higher median level of plasma IL-6 concentrations (1.00 vs. 0.88 pg/ml, P = 0.011). Multivariable models show LV ejection fraction negatively associated with HIV RNA levels [ß -0.18; 95% confidence interval (CI) -0.33, -0.03] and ZDV exposure (ß -1.75; 95% CI -2.62, -0.88) and positively associated with proportion of life on combination antiretroviral treatment (ß 2.65; 95% CI 0.90, 4.41). Higher myocardial performance index was positively associated with serum inflammation marker (IL-6 ß 0.01; 95% CI 0.0001, 0.001). Left ventricular global longitudinal strain was not significantly associated with clinical and laboratory variables of interest. CONCLUSION: Over one-quarter of children and young adults living with HIV demonstrated evidence of cardiac dysfunction, which may be associated with increasing levels of systemic inflammation.

Reduced Transplacental Transfer of Antimalarial Antibodies in Kenyan HIV-Exposed Uninfected Infants.

BACKGROUND: Altered neonatal immune responses may contribute to the increased morbidity observed in HIV-exposed but uninfected (HEU) infants compared with HIV-unexposed uninfected (HUU) infants. We sought to examine the effects of prenatal HIV and malaria exposure on maternal and neonatal plasma cytokine profiles and transplacental antibody transfer. METHODS: Forty-nine HIV+ and 50 HIV- women and their HIV-uninfected neonate pairs from Kenya were assessed. All HIV+ mothers received combination antiretroviral therapy. Maternal plasma and cord blood plasma samples at delivery were tested for 12 cytokines, total IgG, and IgG specific to 4 vaccine antigens and 14 Plasmodium falciparum antigens. RESULTS: HIV+ mothers had lower levels of all 12 plasma cytokines at delivery compared with HIV- mothers, but there were no differences between HEU and HUU neonates. There were no differences in the cord-to-maternal ratios (CMRs) of vaccine-specific IgG between HIV+/HEU and HIV-/HUU maternal-neonate pairs. HIV+/HEU maternal-neonate pairs had significantly lower CMRs for 3 antimalarial IgGs-merozoite surface protein 9, circumsporozoite protein, and erythrocyte binding antigen 181-which remained statistically significant after adjustment for malaria in pregnancy. CONCLUSIONS: In a cohort of optimally treated HIV-infected pregnant women, maternal HIV infection was associated with reduced transplacental transfer of antimalarial antibodies.

Epstein-Barr Virus Type 2 Infects T Cells in Healthy Kenyan Children.

Background: The 2 strains of Epstein-Barr virus (EBV), EBV type 1 (EBV-1) and EBV-2, differ in latency genes, suggesting that they use distinct mechanisms to establish latency. We previously reported that EBV-2 infects T cells in vitro. In this study, we tested the possibility that EBV-2 infects T cells in vivo. Methods: Purified T-cell fractions isolated from children positive for EBV-1 or EBV-2 and their mothers were examined for the presence of EBV and for EBV type. Results: We detected EBV-2 in all T-cell samples obtained from EBV-2-infected children at 12 months of age, with some children retaining EBV-2-positive T cells through 24 months of age, suggesting that EBV-2 persists in T cells. We were unable to detect EBV-2 in T-cell samples from mothers but could detect EBV-2 in samples of their breast milk and saliva. Conclusions: These data suggest that EBV-2 uses T cells as an additional latency reservoir but that, over time, the frequency of infected T cells may drop below detectable levels. Alternatively, EBV-2 may establish a prolonged transient infection in the T-cell compartment. Collectively, these novel findings demonstrate that EBV-2 infects T cells in vivo and suggest EBV-2 may use the T-cell compartment to establish latency.

Maternal Vitamin D Status and Adverse Birth Outcomes in Children from Rural Western Kenya.

Maternal plasma 25-hydroxyvitamin D (25(OH)D) status and its association with pregnancy outcomes in malaria holoendemic regions of sub-Saharan Africa is poorly defined. We examined this association and any potential interaction with malaria and helminth infections in an ongoing pregnancy cohort study in Kenya. The association of maternal plasma 25(OH)D status with pregnancy outcomes and infant anthropometric measurements at birth was determined in a subset of women (n = 63). Binomial and linear regression analyses were used to examine associations between maternal plasma 25(OH)D and adverse pregnancy outcomes. Fifty-one percent of the women had insufficient (<75 nmol/L) and 21% had deficient (<50 nmol/L) plasma 25(OH)D concentration at enrollment. At birth, 74.4% of the infants had insufficient and 30% had deficient plasma 25(OH)D concentrations, measured in cord blood. Multivariate analysis controlling for maternal age and body mass index (BMI) at enrollment and gestational age at delivery found that deficient plasma 25(OH)D levels were associated with a four-fold higher risk of stunting in neonates (p = 0.04). These findings add to the existing literature about vitamin D and its association with linear growth in resource-limited settings, though randomized clinical trials are needed to establish causation.

Reduced Transplacental Transfer of a Subset of Epstein-Barr Virus-Specific Antibodies to Neonates of Mothers Infected with Plasmodium falciparum Malaria during Pregnancy.

Over 35% of children in a region of malaria endemicity are infected with Epstein-Barr virus (EBV) by 6 months of age. This susceptibility may be linked to impaired transplacental transfer of antibodies. In this study, we determined the effect of malaria exposure during pregnancy on the transfer of EBV-specific maternal antibodies in a region of western Kenya that experiences endemic malaria. Pregnant mothers were recruited and followed up until delivery to determine levels of neonatal malaria exposure. Levels of EBV lytic (viral capsid antigen [VCA], Z transcriptional activator [Zta], and early diffuse antigen complex [EAd]) and EBV latent (EBV nuclear antigen-1 (EBNA1]) and tetanus-specific IgG antibodies were measured in 70 paired maternal and cord blood samples using a Luminex-bead-based assay. A high proportion (63%) of the infants were exposed to malaria in utero. Levels of EBV- and tetanus-specific antibodies were similar in malaria-infected mothers and in mothers who had no detectable malaria infection. Malaria-exposed neonates had significantly lower levels of anti-EBNA1, anti-Zta, and anti-EAd antibodies than were seen in their mothers. In utero malaria exposure resulted in significant reductions in transplacental transfer of anti-VCA-p18 and anti-EBNA1 antibodies of 13% and 22%, respectively. Neonates received significantly low levels of anti-Zta and anti-EAd antibodies irrespective of malaria exposure levels. In multivariate analysis, in utero malaria exposure was associated with a significant reduction in the transfer of anti-VCA-p18 and anti-EBNA1 antibodies to the neonates (P = 0.0234 and P = 0.0017, respectively). Malaria during pregnancy results in differential levels of transfer of EBV-specific antibodies from the mother to the fetus. The impaired transplacental transfer of some antibodies may lead to the malaria-exposed neonates being susceptible to early EBV infection.

Epstein-Barr virus genetic variation in lymphoblastoid cell lines derived from Kenyan pediatric population.

Epstein-Barr virus (EBV) is associated with Burkitt's lymphoma (BL), and in regions of sub-Saharan Africa where endemic BL is common, both the EBV Type 1 (EBV-1) and EBV Type 2 strains (EBV-2) are found. Little is known about genetic variation of EBV strains in areas of sub-Saharan Africa. In the present study, spontaneous lymphoblastoid cell lines (LCLs) were generated from samples obtained from Kenya. Polymerase chain reaction (PCR) amplification of the EBV genome was done using multiple primers and sequenced by next-generation sequencing (NGS). Phylogenetic analyses against the published EBV-1 and EBV-2 strains indicated that one sample, LCL10 was closely related to EBV-2, while the remaining 3 LCL samples were more closely related to EBV-1. Moreover, single nucleotide polymorphism (SNP) analyses showed clustering of LCL variants. We further show by analysis of EBNA-1, BLLF1, BPLF1, and BRRF2 that latent genes are less conserved than lytic genes in these LCLs from a single geographic region. In this study we have shown that NGS is highly useful for deciphering detailed inter and intra-variations in EBV genomes and that within a geographic region different EBV genetic variations can co-exist, the implications of which warrant further investigation. The findings will enhance our understanding of potential pathogenic variants critical to the development and maintenance of EBV-associated malignancies.

Breast Milk as a Potential Source of Epstein-Barr Virus Transmission Among Infants Living in a Malaria-Endemic Region of Kenya.

BACKGROUND: We previously reported that infants in Kenya were infected with Epstein-Barr virus (EBV) at <6 months of age, suggesting that mothers were the likely source of transmissible virus to the infant. In this study, we investigated whether breast milk contained infectious EBV and the role of malaria in EBV shedding in breast milk. METHODS: Breast milk samples were obtained from Kenyan mothers at postpartum weeks 6, 10, 14, and 18 and analyzed for presence of infectious EBV. RESULTS: We found that the prevalence of EBV DNA and the mean EBV load were significantly higher at 6 weeks and decreased through postpartum week 18 (P < .0001). High EBV load in breast milk correlated with mothers who had Plasmodium falciparum malaria at delivery. To determine whether viral DNA was encapsidated, breast milk samples were treated with DNAse before DNA extraction. Sixty percent of samples were DNAse resistant, suggesting that the viral DNA in breast milk was encapsidated. Next, we exposed peripheral blood mononuclear cells to breast milk supernatant, which resulted in the generation of EBV-positive lymphoblastoid cell lines, indicating that the virus in breast milk was infectious. CONCLUSIONS: Our data suggest that breast milk contains infectious EBV and is a potential source of viral transmission to infants living in malaria-endemic regions.

Plasmodium falciparum infection is associated with Epstein-Barr virus reactivation in pregnant women living in malaria holoendemic area of Western Kenya.

The role of Plasmodium falciparum malaria in Epstein-Barr virus (EBV) transmission among infants early in life remain elusive. We hypothesized that infection with malaria during pregnancy could cause EBV reactivation leading to high EBV load in circulation, which could subsequently enhance early age of EBV infection. Pregnant women in Kisumu, where P. falciparum malaria is holoendemic, were actively followed monthly through antenatal visits (up to 4 per mother) and delivery. Using real-time quantitative (Q)-PCR, we quantified and compared EBV and P. falciparum DNA levels in the blood of pregnant women with and without P. falciparum malaria. Pregnant women that had malaria detected during pregnancy were more likely to have detectable EBV DNA than pregnant women who had no evidence of malaria infection during pregnancy (64 vs. 36 %, p = 0.01). EBV load as analyzed by quantifying area under the longitudinal observation curve (AUC) was significantly higher in pregnant women with P. falciparum malaria than in women without evidence of malaria infection (p = 0.01) regardless of gestational age of pregnancy. Increase in malaria load correlated with increase in EBV load (p < 0.0001). EBV load was higher in third trimester (p = 0.04) than first and second trimester of pregnancy independent of known infections. Significantly higher frequency and elevated EBV loads were found in pregnant women with malaria than in women without evidence of P. falciparum infection during pregnancy. The loss of control of EBV latency following P. falciparum infection during pregnancy and subsequent increase in EBV load in circulation could contribute to enhanced shedding of EBV in maternal saliva and breast milk postpartum, but further studies are needed.